Molecular characterization of emerging Echovirus 11 (E11) shed light on the recombinant origin of a variant associated with severe hepatitis in neonates.

Echovirus 11 (E11) has gained attention owing to its association with severe neonatal infections. Due to the limited data available, the World Health Organization (WHO) considers public health risk to the general population to be low. The present study investigated the genetic variation and molecular evolution of E11 genomes collected from May to December 2023. Whole genome sequencing (WGS) was performed for 16 E11 strains. Phylogenetic analysis on WG showed how all Italian strains belonged to genogroup D5, similarly to other E11 strains recently reported in France and Germany all together aggregated into separate clusters. A cluster-specific recombination pattern was also identified using phylogenetic analysis of different genome regions. Echovirus 6 was identified as the major recombinant virus in 3Cpro and 3Dpol regions. The molecular clock analysis revealed that the recombination event probably occurred in June 2018 (95% HPD interval: Jan 2016-Jan 2020). Shannon entropy analyses, within P1 region, showed how 11 amino acids exhibited relatively high entropy. Five of them were exposed on the canyon region which is responsible for receptor binding with the neonatal Fc receptor. The present study showed the recombinant origin of a new lineage of E11 associated with severe neonatal infections.

An outbreak of nosocomial infection of neonatal aseptic meningitis caused by echovirus 18.

We describe an outbreak of echovirus 18 infection involving 10 patients in our neonatal intensive care unit (an attack rate of 33%). The mean age at the onset of illness was 26.8 days. Eighty percent were preterm infants. All were discharged home without sequelae. There were no differences in gestation age, birth weight, delivery mode, use of antibiotics, and parenteral nutrition between the enterovirus (EV) group and non-EV group, but the rate of breastfeeding was significantly higher in the EV group. Separation care and reinforcement of hand-washing seemed to be effective in preventing further spread of the virus. Visiting policy, hygiene practice, and handling of expressed breastmilk should be reinforced.

Fulminant echovirus 11 hepatitis in male non-identical twins in northern Italy, April 2023.

Echovirus 11 (E11) has recently been associated with a series of nine neonatal cases of severe hepatitis in France. Here, we present severe hepatitis caused by E11 in a pair of twins. In one of the neonates, the clinical picture evolved to fulminant hepatitis. The E11 genome showed 99% nucleotide identity with E11 strains reported in the cases in France. Rapid genome characterisation using next generation sequencing is essential to identify new and more pathogenetic variants.

Severe and fatal neonatal infections linked to a new variant of echovirus 11, France, July 2022 to April 2023.

We report nine severe neonatal infections caused by a new variant of echovirus 11. All were male, eight were twins. At illness onset, they were 3-5 days-old and had severe sepsis and liver failure. This new variant, detected in France since April 2022, is still circulating and has caused more fatal neonatal enterovirus infections in 2022 and 2023 (8/496; 1.6%, seven associated with echovirus 11) compared with 2016 to 2021 (7/1,774; 0.4%). National and international alerts are warranted.

Temporal phylogeny and molecular characterization of echovirus 30 associated with aseptic meningitis outbreaks in China.

BACKGROUND: An outbreak of aseptic meningitis occurred from June to August 2016, in Inner Mongolia Autonomous Region, China. METHODS: To determine its epidemiological characteristics, etiologic agent, and possible origin, specimens were collected for virus isolation and identification, followed by molecular epidemiological analysis. RESULTS: A total of 363 patients were clinically diagnosed from June 1st to August 31st 2016, and most cases (63.1%, n = 229) were identified between June 22nd and July 17th, with children aged 6 to 12 years constituting the highest percentage (68.9%, n = 250). All viral isolates from this study belonged to genotype C of echovirus 30 (E30), which dominated transmission in China. To date, two E30 transmission lineages have been identified in China, of which Lineage 2 was predominant. We observed fluctuant progress of E30 genetic diversity, with Lineage 2 contributing to increased genetic diversity after 2002, whereas Lineage 1 was significant for the genetic diversity of E30 before 2002. CONCLUSIONS: We identified the epidemiological and etiological causes of an aseptic meningitis outbreak in Inner Mongolia in 2016, and found that Lineage 2 played an important role in recent outbreaks. Moreover, we found that Gansu province could play an important role in E30 spread and might be a possible origin site. Furthermore, Fujian, Shandong, Taiwan, and Zhejiang provinces also demonstrated significant involvement in E30 evolution and persistence over time in China.

Direct next-generation sequencing diagnosis of echovirus 9 meningitis, France.

The prognosis of central nervous system infections caused by enteroviruses partially depends on the viral genotype, which is not provided by current point-of-care diagnostic methods. In this study, next-generation sequencing identified an echovirus 9 directly from the cerebrospinal fluid of a patient presenting with meningitis.

Molecular epidemiological characteristics of echovirus 6 in mainland China: extensive circulation of genotype F from 2007 to 2018.

Echovirus 6 (E6) is associated with various clinical diseases and is frequently detected in environmental sewage. Despite its high prevalence in humans and the environment, little is known about its molecular phylogeography in mainland China. In this study, 114 of 21,539 (0.53%) clinical specimens from hand, foot, and mouth disease (HFMD) cases collected between 2007 and 2018 were positive for E6. The complete VP1 sequences of 87 representative E6 strains, including 24 strains from this study, were used to investigate the evolutionary genetic characteristics and geographical spread of E6 strains. Phylogenetic analysis based on VP1 nucleotide sequence divergence showed that, globally, E6 strains can be grouped into six genotypes, designated A to F. Chinese E6 strains collected between 1988 and 2018 were found to belong to genotypes C, E, and F, with genotype F being predominant from 2007 to 2018. There was no significant difference in the geographical distribution of each genotype. The evolutionary rate of E6 was estimated to be 3.631 × 10-3 substitutions site-1 year-1 (95% highest posterior density [HPD]: 3.2406 × 10-3-4.031 × 10-3 substitutions site-1 year-1) by Bayesian MCMC analysis. The most recent common ancestor of the E6 genotypes was traced back to 1863, whereas their common ancestor in China was traced back to around 1962. A small genetic shift was detected in the Chinese E6 population size in 2009 according to Bayesian skyline analysis, which indicated that there might have been an epidemic around that year.

Echovirus type 6 transmission clusters and the role of environmental surveillance in early warning, the Netherlands, 2007 to 2016.

BackgroundIn the Netherlands, echovirus type 6 (E6) is identified through clinical and environmental enterovirus surveillance (CEVS and EEVS). AimWe aimed to identify E6 transmission clusters and to assess the role of EEVS in surveillance and early warning of E6. MethodsWe included all E6 strains from CEVS and EEVS from 2007 through 2016. CEVS samples were from patients with enterovirus illness. EEVS samples came from sewage water at pre-specified sampling points. E6 strains were defined by partial VP1 sequence, month and 4-digit postcode. Phylogenetic E6 clusters were detected using pairwise genetic distances. We identified transmission clusters using a combined pairwise distance in time, place and phylogeny dimensions. ResultsE6 was identified in 157 of 3,506 CEVS clinical episodes and 92 of 1,067 EEVS samples. Increased E6 circulation was observed in 2009 and from 2014 onwards. Eight phylogenetic clusters were identified; five included both CEVS and EEVS strains. Among these, identification in EEVS did not consistently precede CEVS. One phylogenetic cluster was dominant until 2014, but genetic diversity increased thereafter. Of 14 identified transmission clusters, six included both EEVS and CEVS; in two of them, EEVS identification preceded CEVS identification. Transmission clusters were consistent with phylogenetic clusters, and with previous outbreak reports. ConclusionAlgorithms using combined time-place-phylogeny data allowed identification of clusters not detected by any of these variables alone. EEVS identified strains circulating in the population, but EEVS samples did not systematically precede clinical case surveillance, limiting EEVS usefulness for early warning in a context where E6 is endemic.

Upsurge in echovirus 30 detections in five EU/EEA countries, April to September, 2018.

An upsurge in Echovirus 30 (E30) infections, associated with meningitis/meningoencephalitis, has been observed in Denmark, Germany, the Netherlands, Norway and Sweden in the period April to September 2018, compared with 2015-2017. In total, 658 E30 infections among 4,537 enterovirus infections were detected in 15 countries between January and September 2018 and affected mainly newborns and 26-45 year-olds. National public health institutes are reminded to remain vigilant and inform clinicians of the ongoing epidemic.

Human Rhinoviruses A9, A49, B14 and Echovirus 3, 9 among the patients with acute respiratory infection.

  Background: Acute respiratory infection result in high mortality and morbidity worldwide. There are several viral factors that originate respiratory diseases among them Enteroviruses(EVs) and Human Rhinoviruses(HRVs) can be mentioned. HRVs and EVs belong to Picornaviridae family and they have been recently classified under Enteroviruses. The pattern of respiratory infections generating organisms varies according to geographical locations. Therefore, it seems necessary to organize an appropriate plan to manage common viral diseases exclusively about Rhinoviruses and Enteroviruses. PATIENT AND METHODS: A total of 100 samples were collected from patients with acute respiratory infections (ARIs) who were hospitalized in Ahvaz city hospitals during December 2012 to November 2013 (one year longitude). Semi-Nested PCR was done on samples for detection of HRVs and EVs using region gene of VP4/VP2. Phylogenetic and molecular evolutionary analyses performed with MEGA version 5 software find out the sequence homology among the detected HRV and EV serotype. RESULTS: The results of this study revealed that from of 100 cases of ARIs 19 patients (19%) were HRV positive and 3 (3%) patients positive for EVs. Most positive cases of HRVs were observed in the autumn season while 3 positive cases of EVs were equally found in spring, summer and autumn. Phylogenetic analyses showed that the HRV strains were HRV-A9, HRV-A49, HRV-B14 and EV strains were Echo3 and 9. CONCLUSION: The results of this study revealed that high prevalence of 19% HRVs, HRV-A9, HRV-A49, HRV-B14 serotypes and low frequency of 3% Echo Viruses, Echo3 and Echo 9 serotypes have been detected in patients with ARI.

Molecular characterization of environmental and clinical echovirus 6 isolates from Poland, 2006-2014.

The aim of this study was to investigate the genetic variability of echovirus 6 (E6) isolates from environmental samples and clinical cases of aseptic meningitis from 2006 to 2014. The analysis of the VP1 region showed the extensive diversity (up to 18.8%) and revealed that E6 circulating in Poland belong to four groups. Environmental strains clustered in three groups excepting the 2012 outbreak group, which shows the sudden introduction of new epidemic variant with Asiatic origin. Data from the study established relationships of E6 from Poland with previously characterized strains and confirmed the importance of both clinical and environmental surveillance. J. Med. Virol. 89:936-940, 2017. © 2016 Wiley Periodicals, Inc.

Molecular evolution of two asymptomatic echovirus 6 strains that constitute a novel branch of recently epidemic echovirus 6 in China.

BACKGROUND: Echovirus 6 (E6) infections are associated with aseptic meningitis and acute flaccid paralysis (AFP). But some infections, sometimes most of them, are asymptomatic. The mechanism of E6 virulence is unknown. Analyses of the molecular evolution of asymptomatic E6 may help understand why the infections show different manifestations. METHODS: Ninety-six stool samples of healthy children in Yunnan, China were collected and two E6 strains were isolated from them. The whole genomes of these two E6 strains were sequenced, and their molecular evolution was analyzed. RESULTS: The results showed that the two E6 strains may be derived from KJ7724XX strains, which were predominant in AFP patients in Shangdong in 2011. The evolution was accelerated when the two E6 strains formed, although no positive selection site was found. The 11 exclusive mutations on which selection force significantly changed were found in the 2C, 3AB and 3C genes. CONCLUSION: There are some E6 strains which did not cause the disease in the children of Yunnan. These E6 strains maybe come from a recombinant E6 strain which was associated with the outbreak of AFP in Shangdong in 2011. However, some new mutations were found in the 2C, 3AB and 3C genes of these asymptomatic strains, and these mutations may be constraint by the natural selection and could be potentially responsible for clinical presentations.

Analysis of an echovirus 18 outbreak in Thuringia, Germany: insights into the molecular epidemiology and evolution of several enterovirus species B members.

In October and November 2010, six children and one woman were presented with symptoms of aseptic meningitis in Jena, Thuringia, Germany. Enterovirus RNA was detected in the cerebrospinal fluid of all patients by RT-PCR, and preliminary molecular typing revealed echovirus 18 (E-18) as causative agent. Virus isolates were obtained from stool samples of three patients and several contact persons. Again, most isolates were typed as E-18. In addition, coxsackievirus B5 (CV-B5) and echovirus 25 (E-25) were found to co-circulate. As only few complete E-18 sequences are available in GenBank, the entire genomes of these isolates were determined using direct RNA-sequencing technology. We did not find evidence for recombination between E-18, E-25 or CV-B5 during the outbreak. Viral protein 1 gene sequences and the cognate 3D polymerase gene sequences of each isolate and GenBank sequences were analysed in order to define type-specific recombination groups (recogroups).

Increase in ECHOvirus 6 infections associated with neurological symptoms in the Netherlands, June to August 2016.

The Dutch virus-typing network VIRO-TypeNed reported an increase in ECHOvirus 6 (E-6) infections with neurological symptoms in the Netherlands between June and August 2016. Of the 31 cases detected from January through August 2016, 15 presented with neurological symptoms. Ten of 15 neurological cases were detected in the same province and the identified viruses were genetically related. This report is to alert medical and public health professionals of the circulation of E-6 associated with neurological symptoms.

Elucidation of echovirus 30's origin and transmission during the 2012 aseptic meningitis outbreak in Guangdong, China, through continuing environmental surveillance.

An aseptic meningitis outbreak occurred in Luoding City of Guangdong, China, in 2012, and echovirus type 30 (ECHO30) was identified as the major causative pathogen. Environmental surveillance indicated that ECHO30 was detected in the sewage of a neighboring city, Guangzhou, from 2010 to 2012 and also in Luoding City sewage samples (6/43, 14%) collected after the outbreak. In order to track the potential origin of the outbreak viral strains, we sequenced the VP1 genes of 29 viral strains from clinical patients and environmental samples. Sequence alignments and phylogenetic analyses based on VP1 gene sequences revealed that virus strains isolated from the sewage of Guangzhou and Luoding cities matched well the clinical strains from the outbreak, with high nucleotide sequence similarity (98.5% to 100%) and similar cluster distribution. Five ECHO30 clinical strains were clustered with the Guangdong environmental strains but diverged from strains from other regions, suggesting that this subcluster of viruses most likely originated from the circulating virus in Guangdong rather than having been more recently imported from other regions. These findings underscore the importance of long-term, continuous environmental surveillance and genetic analysis to monitor circulating enteroviruses.

Acute meningoencephalitis associated with echovirus 9 infection in Sri Lanka, 2009.

The aetiology of acute meningoencephalitis in Sri Lankan children and adults is poorly understood. This study was carried out to determine pathogens responsible for meningoencephalitis in Sri Lanka. A hospital-based cross-sectional study was performed using cerebrospinal fluid samples (22 adult and 17 pediatric) collected from August to December 2009 from patients clinically diagnosed with acute meningoencephalitis at two tertiary care hospitals in Sri Lanka. Routine microbiology for bacterial pathogens together with in-house RT-PCR and PCR assays for the detection of dengue viruses, Japanese encephalitis virus, West Nile virus, chikungunya virus, enteroviruses, mumps virus, measles virus, herpes simplex viruses types 1 and 2, and varicella zoster virus were performed. Bacterial pathogens were not isolated from any patient specimens. However, from nine of the paediatric patients aged 1 month to 10 years (mean age 5.2 years) echovirus 9 (E-9; family Picornaviridae, genus Enterovirus,species Enterovirus B ) was detected by RT-PCR. All nine patients presented with fever, six had headache, and seven had vomiting. Neck stiffness indicating meningitis was present in six of the patients. Phylogenetic analysis of partial VP1 and VP4-VP2 genes showed these E-9 strains to be most closely related to E-9 strains detected in CSF from Korea and France in 2005 and 2006. The remaining patients were negative for all other viruses tested. E-9 was the most common cause of acute meningoencephalitis in the tested paediatric population from Sri Lanka in 2009, which likely reflects circulation of this E-9 strain between Europe and Asia over several years.

Aseptic meningitis outbreak associated with echovirus 30 among high school football players--Los Angeles County, California, 2014.

On August 4, 2014, the Acute Communicable Disease Control Program of the Los Angeles County Department of Public Health received a report of three aseptic meningitis cases among football players at a county high school. An investigation was conducted to determine the extent of the outbreak, identify potential exposures, and recommend control measures. An outbreak-associated aseptic meningitis case was defined as an illness of any team or family member with onset during July 28-August 11 with 1) cerebrospinal fluid pleocytosis and negative bacterial culture or 2) an emergency department visit with headache, fever, and stiff neck. Ten cases were identified; nine in males, and one in a female; patient ages ranged from 13 to 17 years. All the patients sought care at an emergency department, and five were hospitalized, resulting in 12 total hospital days. All 10 patients have recovered. Eight patients were football players, and two were siblings of football players. The most affected subgroup was the junior varsity football team, with seven cases out of 57 players (attack rate = 12.3%); the relative risk for aseptic meningitis was higher among players who were linemen than among those who were not linemen (relative risk = 5.4 [p = 0.03]). Of the 10 patients, eight tested positive by polymerase chain reaction for enterovirus, and two were not tested. Echovirus testing was performed at the California Viral and Rickettsial Disease Laboratory. Of the eight specimens testing positive for enterovirus, seven tested positive for echovirus 30, and one specimen could not be typed because of insufficient quantity.

Recombination and evolutionary dynamics of human echovirus 6.

Enterovirus (EV) infections are associated with a wide array of often severe disease presentations including aseptic meningitis, encephalitis, and acute flaccid paralysis. Surveillance for polioviruses and other EVs is therefore important as a public health measure both for patient management and epidemiological studies. From 1988 to 2008, echovirus (E) 30 was the predominant genotype in Spain (33.7% of the total typed EVs). E6 was also endemic throughout this period although isolated less frequently (12.5%). In 2009, however, a substantial increase in the incidence of E6 was detected (60%), displacing E30 type (2%). To investigate the evolution and recombination in the epidemiology and transmission of E6 in Spain, a genetic analysis in VP1 and 3Dpol regions of 67 Spanish strains collected during the period 2004-2010 was performed. All VP1 sequences clustered monophyletically in the assigned genogroup C, subgroup 9, currently the predominant circulating strains identified in Europe and elsewhere in the last 10 years. 3Dpol sequences were interspersed with other species B EVs resulting from several recombination events that generated at least 12 different recombinant forms (RFs) among study samples. These showed typically minimal divergence in VP1. The co-circulation of different lineages of E6 in the same geographical area associated with its mainly endemic pattern of transmission may have contributed to the extremely short estimated half-life of E6 RFs (0.87 years). This pattern contrasts markedly with other species B EVs and EV71 where VP1 lineage expansion and extinction occurred in step with defined recombination events and periodic changes in incidence.

Detection and genotyping of enteroviruses in cerebrospinal fluid in patients in Victoria, Australia, 2007-2013.

Genotyping by VP1 fragment polymerase chain reaction (PCR) and nucleic acid sequencing to detect enterovirus (EV) genotypes was performed directly on 729 EV PCR positive cerebrospinal fluid (CSF) samples collected between 2007 and 2012 from Victorian hospital inpatients. The overall genotype identification rate from CSF-positive material was 43%. The four most common genotypes identified were Echovirus 6 (24%), Echovirus 30 (17%), Echovirus 25 (10%), and Coxsackievirus A9 (10%), together comprising 61% of all EVs typed. The seasonal distribution of all EVs identified followed the recognized pattern of mainly summer epidemics. Three of the four predominant genotypes were present in each of the 6 years in which the study was conducted, with 20 other EV genotypes also detected, often in only a single year. Genotyping of EVs directly in CSF is faster, simpler and more sensitive than traditional virus neutralization assays performed on EV positive samples.

Rubella virus genotype 1G and echovirus 9 as etiologic agents of exanthematous diseases in Brazil: insights from phylogenetic analysis.

The aim of the present study was to identify the rubella virus (RV) and enterovirus (EV) genotypes detected during the Epidemiological Surveillance on Exanthematic Febrile Diseases (VIGIFEX) study and to perform phylogenetic analysis. Ten RV- and four EV-positive oropharyngeal samples isolated from cell culture were subjected to RT-PCR and sequencing. Genotype 1G and echovirus 9 (E-9) was identified in RV- and EV-positive samples, respectively. The RV 1G genotype has been persisting in Brazil since 2000-2001. No evidence of E-9 being involved in exanthematic illness in Brazil has been reported previously. Differential laboratory diagnosis is essential for management of rash and fever disease.